Human B7-H3 HEK293 Overexpression Lysate: Product Information
This Human B7-H3 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of B7-H3 protein (Cat: 11188-H08H) from the overexpression lysate was verified.
A DNA sequence encoding the human CD276 (Q5ZPR3-1) extracellular domain (Met 1-Thr 461) was expressed, with a polyhistidine tag at the C-terminus.
The recombinant human CD276 consists of 444 amino acids and predictes a molecular mass of 48 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of rhCD276 is approximately 73 kDa due to glycosylation.
Human B7-H3 HEK293 Overexpression Lysate: Usage Guide
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube.
2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Western Blot (WB) Optimal dilutions/concentrations should be determined by the end user.
Human B7-H3 HEK293 Overexpression Lysate: Alternative Names
Human 4Ig-B7-H3 Overexpression Lysate; Human B7-H3 Overexpression Lysate; Human B7H3 Overexpression Lysate; Human B7RP-2 Overexpression Lysate
B7-H3 Background Information
B7-H3 is a member of the B7 family of immune regulatory ligands that is thought to attenuate peripheral immune responses through co-inhibition. It plays an important role in adaptive immune responses, and was shown to either promote or inhibit T-cell responses in various experimental systems. B7-H3 may play an important role in muscle-immune interactions, providing further evidence of the active role of muscle cells in local immunoregulatory processes. B7-H3 is a novel protein structurally related to the B7 family of ligands by the presence of a single set of immunoglobulin-V-like and immunoglobulin-C-like (VC) domains. Previous studies have correlated its overexpression with poor prognosis and decreased tumor-infiltrating lymphocytes in various carcinomas including uterine endometrioid carcinomas, and mounting evidence supports an immuno-inhibitory role in ovarian cancer prognosis. Recently, B7-H3 expression has been reported in several human cancers indicating an additional function of B7-H3 as a regulator of antitumor immunity.
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